Methods to grow replicative cultures of normal human bronchial epithelial cells have been developed. These cells, which can be subcultured and will undergo 35 population doublings, have the expected epithelial cell characteristics of keratin, desmosomes, blood group antigens and the normal chromosomal karyotype. They form colonies when plated at low cell density. Two percent serum and mouse 3T3 feeder cells are required for growth; addition of EGF, fibronectin and cholera toxin markedly increase the clonal growth rate and cloning efficiency. Explant cultures of human bronchus have been maintained for more than one year. In addition, long-term continuous growth of epithelial cells has been accomplished by repeated transfer of a tissue explant. After 6 months and 13 transfers, epithelial cells continue to grow out from both control and carcinogen treated explants. Human prostatic epithelial cells previously infected with SV40 virus have been repeatedly exposed to 4NQO. The carcinogen treatment caused extensive aneuploidy. The tumorigenic potential of these cells is being assessed in athymic nude mice.